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1 doi:1.138/nature9464 C 3 K 3 -d 8 C 3 PK-d 7 MK-4-d 12 C 3 C 3 MK-4-d 7 C 3 Supplementary Figure 1 Chemical structures of PK-d 7, MK-4-d 12, K 3 -d 8 and MK-4-d 7. WWW NATURE.CM/NATURE 1

2 a Relative ratio (UBIA1/ β-actin) sicontrol siubia1 b Relative ratio (CQ2/ β-actin) sicontrol sicq2 Supplementary Figure 2 UBIA1 and CQ2 mrna expression in sirna-transfected MG-63 cells. a, UBIA1 mrna expression in UBIA1 sirna (siubia1)-transfected MG-63 cells. Values are means ± S.E. (n = 6). *** P <.1 versus control sirna (sicontrol)-transfected cells. b, CQ2 mrna expression in CQ2 sirna (sicq2)-transfected MG-63 cells. Values are means ± S.E. (n = 6). *** P <.1 versus control sirna (sicontrol)- transfected cells. WWW NATURE.CM/NATURE 2

3 a Relative ratio (UBIA1 / β-actin) Mock siubia1 pcna3.3 -UBIA1 b 36.8 ka Mock siubia1 pcna3.3 -UBIA1 Supplementary Figure 3 UBIA1 mrna expression and protein level in siubia1 or UBIA1 expression vector-transfected MG-63 cells. a, Expression of UBIA1 mrna in MG-63 cells: mock transfected and transfected with siubia1 or pcna3.3 UBIA1 expression vector. Values are means ± s.e.m. (n = 6). *** P <.1 versus each cell. b, Western blot analysis of siubia1 or UBIA1 expression in vector-transfected MG-63 cells. WWW NATURE.CM/NATURE 3

4 Sf9-Control Sf9-UBIA1 UBIA1 Supplementary Figure 4 Western blot analysis for UBIA1 expression in UBIA1 baculovirus-infected Sf9 cells. WWW NATURE.CM/NATURE 4

5 Relative ratio (Ubiad1/ β-actin) Seminal vesicle Testis Uterus vary Bone Kidney Spleen Pancreas Liver Small intestine uodenum Stomach Lung Aorta Parathyroid gland Thyroid Thymus Heart culus Medulla oblongata Cerebellum Interbrain Midbrain Cerebrum Supplementary Figure 5 Ubiad1 mrna expression in mouse tissues. Total RNAs of mouse tissues (five female C57BL/6 mice aged 8 weeks) were isolated with Isogen (Nippon Gene) according to the manufacturer s protocol. First strand cna was made with AMV reverse transcriptase (TaKaRa). cna were mixed with a SYBR Green core reagent kit (PE Biosystems), and real-time PCR used a CFX96 real-time PCR system (Bio- Rad). We used mouse Ubiad1 (GenBank accession No. NM_27873, FP: , RP: ) and mouse β-actin (GenBank accession No. X3672, FP:25-271, RP:35-326). WWW NATURE.CM/NATURE 5

6 a 1.8 Relative ratio (Ubiad1/ β-actin) b 7 Cerebrum Cerebellum Heart Liver Pancreas Kidney Bone vary Uterus Concentration ( pmol/g) MK-4-d 7 MK-4-d 7 epoxide 1 Uterus vary Bone Kidney Pancreas Liver Heart Cerebellum Cerebrum c Concentration ( pmol/g) MK-4 MK-4 epoxide 2 Uterus vary Bone Kidney Pancreas Liver Heart Cerebellum Cerebrum Supplementary Figure 6 Ubiad1 mrna expression in mouse tissues and concentrations of MK-4, MK-4 epoxide, MK-4-d 7 and MK-4-d 7 epoxides in tissues of mice administered PK-d 7. a, Ubiad1 mrna expression in mouse tissues. b, Concentrations of MK-4-d 7 and MK- 4-d 7 epoxide in tissues of mice administered PK-d 7. c, Concentrations of MK-4 and WWW NATURE.CM/NATURE 6

7 MK-4 epoxide accumulated in mouse tissues. Five 8-week-old female C57BL/6 mice (SLC, Inc.) in each group were given either vehicle or PK-d 7 orally as a single dose at 1 mmol/kg body weight. After 6 h, mice were sacrificed and tissues were removed and stored at 8 C for analysis. Measurements for MK-4, MK-4 epoxide, MK-4-d 7 and MK-4-d 7 epoxide in tissues used the same LC-APCI-MS/MS methods as described previously 13. (ka) Microsome Sf9- control Sf9- UBIA Supplementary Figure 7 Western blot analysis for UBIA1 expression in microsomes from UBIA1 baculovirus-infected Sf9 cells. WWW NATURE.CM/NATURE 7

8 Sf9-UBIA1 microsome Intensity (cpm) Substrate: PK-d 7 Substrate: K 3 -d e 5 MK-4-d e 5 MK-4-d 7 Intensity (cpm) 1.6 e e e 5 1. e 5 8. e 4 6. e e e Time (min) Time (min) Supplementary Figure 8 LC-APCI-MS/MS analysis of MK-4-d 7 converted from PK-d 7 or K 3 -d 8 in the microsomal fraction of UBIA1 baculovirus-infected Sf9 cells. MRM chromatogram of MK-4-d 7 converted from PK-d 7 or K 3 -d 8 in the microsomal fraction of UBIA1 baculovirus-infected Sf9 cells. WWW NATURE.CM/NATURE 8